The role of c-Fms and GM-Csf in intestinal biology

© 2012 Dr. Dilara AkcoraCsf-1 and GM-CsF are important growth factors required for proliferation, cell fate specification and differentiation of hematopoietic stem cell and progenitors. Their role in immunity, inflammation and progression of both hematopoietic and non-hematopoietic cancers has been extensively investigated. Despite their known role in inflammation and the presence of the receptors for Csf-1 (c-Fms) and GM-Csf (GMCsfR) on small intestinal epithelial cells, there is little evidence for their contribution to the gastrointestinal homeostasis. Recent investigation however confirmed that the complete loss of c-Fms can lead to an alteration of intestinal proliferation and differentiation. However, it was not clear whether this effect was direct or indirectly mediated by tissue macrophages that also express Csf-1 and c-Fms. Furthermore, the role of GM-Csf has not been investigated previously in terms of small intestine and colon homeostasis. In this thesis, I explored the role of c-Fms and GM-Csf in intestinal biology. The first aim of this study was to directly examine the effect of loss of c-Fms in small intestine using organoid cultures and a tamoxifen inducible, adult intestine specific c-fms conditional knock-out mouse model. The expression pattern of c-Fms in SI epithelium was investigated. The intrinsic deficiency of c-Fms produced a reduction in organoid forming efficiency which was associated with down-regulation of intestinal stem cell gene expression at the mRNA level. In vivo loss of c-Fms produced a decrease in Paneth and enteroendocrine cell numbers as well as a positional shift of proliferating cells towards the bottom of crypts previously occupied by Paneth cells. The second aim of this study was to examine the effects of Csf-1 and IL-34 as well as the intestinal mitogens Wnt3a and R-spondin1 on proliferation in immortalized colon cell lines. Furthermore, I explored the hypothesis that CD24+ colon cells provide a niche to colonic stem cells with the underlying rationale that CD24+ Paneth cells are known to contribute to the maintenance of SI stem cells. Finally, the effect of intestine specific loss of c-Fms in adult colon was investigated. It was found that only Csf-1 or R-spondin1 alone can stimulate colon epithelial growth and when combined together this stimulatory effect was further increased. CD24+ colon cells were enriched for goblet cells and a cell population in which Lgr-5 expression was relatively high. The expression of CD24 expression was surprisingly not affected by the loss c-Fms in colon crypts. In contrast to the global c-fms knock-out study, intestine specific inactivation of c-fms led to unimpaired epithelial cell proliferation and differentiation at least over 4 to 8 weeks. The last aim of my thesis was to explore the role of GM-Csf in SI and colon homeostasis through the use of mice lacking of GM-Csf expression. I found that the loss of GM-Csf resulted in a dramatic reduction of intestinal stem cell and proliferation gene expression in gm-csf -/- female mice. Interestingly this was not the case in gm-csf -/- male mice. In addition, intestinal proliferation and differentiation were not altered by the loss of GM-Csf in female mice. These data emphasize the importance of sex related differences in the regulation of intestinal stem cells in mice. Collectively, these data suggest that c-Fms signaling is important for SI homeostasis by mediating Paneth and enteroendocrine cell differentiation and promoting stem cell function. The function of c-Fms might be dispensable for colon epithelial cell differentiation in adult mice or perhaps the absence of c-Fms might be compensated for by other essential factors released by the stroma such as macrophages or myofibroblasts. Moreover, c-Fms activation by Csf-1 may co-operate with Wnt signaling to promote colon cell proliferation. In addition, my work has unraveled a potential role for GM-Csf in sex specific transcriptional regulation of intestinal stem cell and proliferation genes. Overall, my thesis studies have advanced our understanding of the role of c-Fms and GM-Csf in intestinal biology